Stroke is a major cause of morbidity and disability and the functional recovery is slow and uncertain. Our data demonstrate that intravenously transplanted bone marrow stromal cells (MSCs), including mesenchymal stem or progenitor cells which possess multilineage developmental potential, migrate to the damaged brain tissue, and improve functional recovery. This research proposal is concerned with understanding the cellular and molecular mechanisms responsible for the therapeutic effect of MSC therapy of stroke. The major hypothesis being tested in vivo is that the MSCs delivered to the brain after stroke provide neuroprotection by reducing apoptotic cell death of neurons, promote angiogenesis, and induce structural changes in neurons (neuronal remodeling), including axonal and dendritic sprouting and synapse formation in the penumbral region. To determine whether MSCs differentiate into neural cells and integrate into the cerebral tissue and form connections, ultrastructural (electron microscopy) and electrophysiological measurements will be performed on MSCs that have been labeled prior to transplantation with green fluorescent protein (GFP) and gold particles using gene gun technology. We will test the hypothesis that the microenvironment of the ischemic brain tissue induces in MSCs and astrocytes the production of angiogenic and neurotrophic growth factors (VEGF, bFGF, BDNF, NGF), which mediate the beneficial effects of MSC treatment. Using appropriate in vitro model systems, detailed functional characterization of MSC and astrocyte secreted products will be performed to assess the antiapoptotic, angiogenic, and neurogenic activities of these molecules. In addition, we will test whether neurotrophic factors alter the growth and differentiation pathways of MSCs to become neural cells (glia and neurons). Thus, the studies in this proposal will provide insight into the cellular and molecular mechanisms underlying the therapeutic benefit provided by MSC cellular therapy of stroke.